Abstract: The Laser Scanning Confocal Microscopy (LSCM) has a unique capability to resolve below- surface tissue structures at a sub-cellular level. With this capability, LSCM has contributed compelling results in early cancer detection. Conventionally, LSCM is based on an assembly of bulky, expensive components[unreadable]motor-controlled scanning mirrors, translation stages, and high numerical aperture (NA) lenses[unreadable]to achieve sub-cellular resolution and optical sectioning. Consequently, applications of LSCM in cancer diagnosis, especially for cancer in hard-to-reach parts of human bodies (such as digestive and respiratory systems), have been limited. In order to maximize the potential of LSCM in early cancer diagnosis, it is necessary to develop miniature LSCM systems that can be fitted on the tip of an endoscope probe or catheter needle to facilitate high-resolution, minimally-invasive, in vivo confocal microscopic imaging. In view of the tremendous potential and significant challenges in developing miniature LSCM, I propose a transformative technical route-Optofluidic Laser Scanning Confocal Microscope (O- LSCM). Specifically, I will (1) address optical and mechanical challenges in the implementation of O-LSCM, (2) characterize and optimize pre-packaged O-LSCM, and (3) construct O-LSCM imaging probe and apply it to tissue imaging for early cancer detection. The proposed O-LSCM realizes all optical and mechanical functions needed for an LSCM in a single embodiment via the control of fluid flows in a microfluidic chamber. O-LSCM has no moving components, and it can be conveniently realized with a simple micro-fabrication technique (i.e., micro mold injection). Therefore, fabrication and assembly process can be dramatically simplified, the assembled system can be miniaturized to such an extent that it is suitable for endoscopic applications, and reliability and optical alignment precision of the system can be significantly improved. With its strong functionalities, simplicity, compactness, and robustness, the proposed O-LSCM will facilitate the design of next-generation endoscopic confocal microscopy systems and will benefit a wide range of biological studies and clinical applications. Public Health Relevance: Laser scanning confocal microscopy (LSCM) is a powerful technique for obtaining high resolution three-dimensional (3D) microscopic images of various biological samples. The long-term objective of this grant is to invent a paradigm-shifting technical route to LSCM-Optofluidic Laser Scanning Confocal Microscope (O-LSCM). With its strong functionalities, simplicity, compactness, and robustness, the proposed O-LSCM will facilitate the design of next-generation high-resolution, minimum-invasive endoscopic confocal microscopy systems and will benefit a wide range of biological studies and clinical applications (such as early cancer detection).